Helicobacter pylori is a bacterial pathogen that chronically colonizes the human stomach. One of the distinctive features of H. pylori is the lipid A moeity of its lipopolysaccharide (LPS). Compared to the prototypical Enterobacteriaceae lipid A, it has a unique structure and is much reduced in its endotoxic activity. Our hypothesis is that H. pylori LPS has different effects on host cell signaling pathways than Escherichia coli LPS. Further we speculate that these differences result in the inability of the human host to respond and clear the H. pylori infection. We will take a genome-wide approach to detect and analyze changes in the human epithelial cell response to H. pylori LPS. In Specific Aim 1, we will use microarray analysis to detect changes in gene expression in gastric epithelial cells treated with H. pylori LPS compared to untreated cells and those treated with E. coli LPS. Whether these changes are due to the recognition by Toll-like receptor 2 (TLR2) or TLR4 will also be assessed using cells either with transfected TLR2 or TLR4, or with dominant negative versions of these receptors. These findings will be confirmed by RT-PCR. In Specific Aim 2, a proteomic approach will be taken; protein profiles of epithelial cells after H. pylori LPS or E. coli LPS treatment will be compared by 2D gel electrophoresis. Proteins of interest will be identified by mass spectrometric analysis. Together these approaches should allow us to determine the specific program of host responses to H. pylori LPS. This information should give us a better understanding of how this organism is able to chronically persist in the stomach and evade eradication by the host immune system. In Specific Aim 3, we will isolate H. pylori strains with alterations in the lipid A structure to determine which portion is responsible for the reduced endotoxic activity. The long-term goal of this research will be to develop a rational approach for new therapeutic treatments for H. pylori infections. Further we anticipate that deciphering the transcriptional and protein profiles resulting from H. pylori LPS will provide us with a framework to recognize the diagnostic features of chronic infections.